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1.
J Cancer Res Clin Oncol ; 146(4): 897-907, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32146565

RESUMO

INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is a very aggressive cancer. There are various sub-cellular events (both genetic and epigenetic) that get dysregulated leading to tumorigenesis. Methylation in promoters of tumor suppressor genes is one of these epigenetic phenomena contributing to the pathogenesis of cancer. Genes analyzed for promoter methylation status in this study namely SPARC (Secreted Protein Acidic and Rich in Cysteine, UCHL1 (ubiquitin carboxy-terminal hydrolase L1), NPTX2 (neuronal pentraxin 2), PENK (proenkephalin) had been studied in pancreatic cancer, but there is a need to check methylation in these genes as circulatory non-invasive markers. This study analyzed the absolute quantification of methylation levels of SPARC, UCHL1, PENK, and NPTX2 genes promoters in PDAC patients as well as in chronic pancreatitis (CP) patients and healthy subjects (HC) and evaluated its clinical significance in PDAC. MATERIALS AND METHODS: The study included 65 PDAC patients, 25 CP patients, and 25 healthy controls. DNA was extracted from their plasma samples and subsequently given bisulfite treatment. Absolute quantization of methylated and unmethylated copies of gene promoters of all the four genes was performed using real-time PCR (SYBR green) by the standard curve method. Methylation levels were expressed as methylation index (MI) for each gene in each patient. MI was calculated from absolute copy numbers as follows: MI-methylated copy number/methylated copy number + unmethylated copy number). These indices were used to compare gene methylation levels within different groups and to correlate with clinicopathological features and survival of pancreatic cancer patients. An appropriate statistical analysis was applied. RESULTS: Methylation indices for all the four genes in PDAC cases were found to be significantly higher as compared to that in healthy individuals. SPARC MI values were found to differentiate early-stage PDAC patients from CP patients. PDAC patients with the metastasized disease and stage IV disease were found to have high MI for the SPARC gene as well as for the NPTX2 gene, while a higher UCHL1 methylation index was found to correlate with an advanced stage of the disease. Higher MI values for SPARC and NPTX2 genes were found to associate with poor survival in patients with PDAC. CONCLUSION: Methylation load in the form of MI for each of the four genes assessed in plasma may emerge as a non-invasive biomarker to differentiate pancreatic cancer from healthy individuals. But only SPARC and NPTX2 hypermethylation were able to distinguish pancreatic cancer from chronic pancreatitis. Association of aberrant methylation in SPARC and NPTX2 gene with metastasis and poor survival of patients suggest the role of methylation in these genes as prognostic markers.


Assuntos
Carcinoma Ductal Pancreático/genética , DNA Tumoral Circulante/genética , Metilação de DNA , Genes Supressores de Tumor , Neoplasias Pancreáticas/genética , Proteína C-Reativa/genética , Carcinoma Ductal Pancreático/sangue , Estudos de Casos e Controles , Diferenciação Celular/genética , DNA Tumoral Circulante/sangue , Encefalinas/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Osteonectina/genética , Neoplasias Pancreáticas/sangue , Pancreatite Crônica/sangue , Pancreatite Crônica/genética , Regiões Promotoras Genéticas , Precursores de Proteínas/genética , Ubiquitina Tiolesterase/genética
2.
BMC Cancer ; 19(1): 881, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31488082

RESUMO

BACKGROUND: Gene expression profiling of rare cancers has proven challenging due to limited access to patient materials and requirement of intact, non-degraded RNA for next-generation sequencing. We customized a gene expression panel compatible with degraded RNA from formalin-fixed, paraffin-embedded (FFPE) patient cancer samples and investigated its utility in pathway activity profiling in patients with metaplastic breast cancer (MpBC). METHODS: Activity of various biological pathways was profiled in samples from nineteen patients with MpBC and 8 patients with invasive ductal carcinoma with triple negative breast cancer (TNBC) phenotype using a custom gene expression-based assay of 345 genes. RESULTS: MpBC samples of mesenchymal (chondroid and/or osteoid) histology demonstrated increased SNAI1 and BCL2L11 pathway activity compared to samples with non-mesenchymal histology. Additionally, late cornified envelope and keratinization genes were downregulated in MpBC compared to TNBC, and epithelial-to-mesenchymal transition (EMT) and collagen genes were upregulated in MpBC. Patients with high activity of an invasiveness gene expression signature, as well as high expression of the mesenchymal marker and extracellular matrix glycoprotein gene SPARC, experienced worse outcomes than those with low invasiveness activity and low SPARC expression. CONCLUSIONS: This study demonstrates the utility of gene expression profiling of metaplastic breast cancer FFPE samples with a custom counts-based assay. Gene expression patterns identified by this assay suggest that, although often histologically triple negative, patients with MpBC have distinct pathway activation compared to patients with invasive ductal TNBC. Incorporation of targeted therapies may lead to improved outcome for MpBC patients, especially in those patients expressing increased activity of invasiveness pathways.


Assuntos
Carcinoma Ductal de Mama/genética , Receptores de Fatores de Crescimento/metabolismo , Transdução de Sinais/genética , Transcriptoma/genética , Neoplasias de Mama Triplo Negativas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteína 11 Semelhante a Bcl-2/metabolismo , Carcinoma Ductal de Mama/patologia , Estudos de Coortes , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Osteonectina/genética , Fenótipo , Prognóstico , Fatores de Transcrição da Família Snail/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
3.
Exp Eye Res ; 188: 107787, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31479653

RESUMO

Cataract-associated gene discovery in human and animal models have informed on key aspects of human lens development, homeostasis and pathology. Additionally, in vitro models such as the culture of permanent human lens epithelium-derived cell lines (LECs) have also been utilized to understand the molecular biology of lens cells. However, these resources remain uncharacterized, specifically regarding their global gene expression and suitability to model lens cell biology. Therefore, we sought to molecularly characterize gene expression in the human LEC, SRA01/04, which is commonly used in lens studies. We first performed short tandem repeat (STR) analysis and validated SRA01/04 LEC for its human origin, as recommended by the eye research community. Next, we used Illumina HumanHT-12 v3.0 Expression BeadChip arrays to gain insights into the global gene expression profile of SRA01/04. Comparative analysis of SRA01/04 microarray data was performed using other resources such as the lens expression database iSyTE (integrated Systems Tool for Eye gene discovery), the cataract gene database Cat-Map and the published lens literature. This analysis showed that SRA01/04 significantly expresses >40% of the top iSyTE lens-enriched genes (313 out of 749) across different developmental stages. Further, SRA01/04 also significantly expresses ~53% (168 out of 318) of cataract-associated genes in Cat-Map. We also performed comparative gene expression analysis between SRA01/04 cells and the previously validated mouse LEC 21EM15. To gain insight into whether SRA01/04 reflects epithelial or fiber cell characteristics, we compared its gene expression profile to previously reported differentially expressed genes in isolated mouse lens epithelial and fiber cells. This analysis suggests that SRA01/04 has reduced expression of several fiber cell-enriched genes. In agreement with these findings, cell culture analysis demonstrates that SRA01/04 has reduced potential to initiate spontaneous lentoid body formation compared to 21EM15 cells. Next, to independently validate SRA01/04 microarray gene expression, we subjected several candidate genes to RT-PCR and RT-qPCR assays. This analysis demonstrates that SRA01/04 supports expression of many key genes associated with lens development and cataract, including CRYAB, CRYBB2, CRYGS, DKK3, EPHA2, ETV5, GJA1, HSPB1, INPPL1, ITGB1, PAX6, PVRL3, SFRP1, SPARC, TDRD7, and VIM, among others, and therefore can be relevant for understanding the mechanistic basis of these factors. At the same time, SRA01/04 cells do not exhibit robust expression of several genes known to be important to lens biology and cataract such as ALDH1A1, COL4A6, CP, CRYBA4, FOXE3, HMX1, HSF4, MAF, MEIS1, PITX3, PRX, SIX3, and TRPM3, among many others. Therefore, the present study offers a rich transcript-level resource for case-by-case evaluation of the potential advantages and limitations of SRA01/04 cells prior to their use in downstream investigations. In sum, these data show that the human LEC, SRA01/04, exhibits lens epithelial cell-like character reflected in the expression of several lens-enriched and cataract-associated genes, and therefore can be considered as a useful in vitro resource when combined with in vivo studies to gain insight into specific aspects of human lens epithelial cells.


Assuntos
Biomarcadores , Células Epiteliais/citologia , Proteínas do Olho/genética , Cristalino/citologia , Linhagem Celular , Efrina-A2/genética , Células Epiteliais/metabolismo , Expressão Gênica/fisiologia , Humanos , Cristalino/metabolismo , Biologia Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Osteonectina/genética , Fator de Transcrição PAX6/genética , Reação em Cadeia da Polimerase em Tempo Real , Ribonucleoproteínas/genética , Cadeia B de alfa-Cristalina/genética , Cadeia B de beta-Cristalina/genética
4.
Life Sci ; 229: 277-287, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31150687

RESUMO

AIMS: Secreted protein acidic and rich in cysteine, (SPARC), is a matricellular protein implicated in the modulation of the extracellular matrix (ECM) and mitochondrial proteins expression. MAIN METHODS: To study the mechanism through which SPARC is involved in the possible link between ECM and mitochondria, C2C12 myoblasts were cultured with/without the exogenous addition/inhibition of SPARC as well as activation/inhibition of adenosine monophosphate-activated protein kinase (AMPK). Electrical pulse stimulation (EPS), was applied for 2 days in myotubes. KEY FINDINGS: The expressions of ECM-related (integrin-linked kinase (ILK), glycogen synthase kinase-3 beta (GSK-3ß), phosphorylated-GSK-3ß (p-GSK-3ß) and collagen 1a1), mitochondrial-related (AMPK, phosphorylated-AMPK (p-AMPK), succinate dehydrogenase (SDHB) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (Pgc1α)) and SPARC proteins and/or genes were measured after modulation of SPARC and/or AMPK as well as with or without EPS. The addition of SPARC in C2C12 myoblast increased the expression of ILK, p-GSK-3ß and p-AMPK whereas anti-SPARC antibody decreased them at different incubation times (0, 10, and 30 min, and 6 h). The AMPK activation increased SPARC, collagen 1a1, p-AMPK and SDHB proteins level, however, AMPK inhibition blunted the effects. EPS induced Sparc and Pgc1a genes expression. SIGNIFICANCE: Sparc, an EPS-induced gene, may be involved in the link between ECM remodeling and mitochondrial function in muscle via its interaction with ILK/AMPK.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Matriz Extracelular/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Mioblastos/metabolismo , Osteonectina/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Células Cultivadas , Estimulação Elétrica , Regulação da Expressão Gênica , Camundongos , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Mioblastos/citologia , Osteonectina/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fosforilação , Proteínas Serina-Treonina Quinases/genética
5.
Exp Mol Pathol ; 110: 104276, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31233732

RESUMO

Secreted protein acidic and rich in cysteine (SPARC) plays a crucial role in the malignant progression of a number of human cancers. However, the roles of SPARC in lung squamous cell carcinoma (LSCC) remain elusive. In this present study, we first detected SPARC expression and investigated the relationship between SPARC expression and the clinicopathological attributes of LSCC patients. Then we constructed SPARC-overexpression model in LSCC cell line to explore the characteristics of SPARC in LSCC development both in vitro and in vivo. The data demonstrated a remarkably higher level of SPARC in LSCC tissues than in corresponding non-cancerous tissues and elevated SPARC expression was significantly correlated with poor outcome in LSCC patients. Moreover, a serial of phenotypic experiments indicated that SPARC overexpression substantially facilitated the growth and inhibited the apoptosis in LSCC cells and xenografts. Taken together, our results suggest that SPARC is a novel prognostic marker for LSCC prognosis and SPARC significantly promotes LSCC tumorigenesis. Targeting SPARC may provide a novel therapeutic strategy for LSCC management.


Assuntos
Carcinogênese/genética , Carcinoma de Células Escamosas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Osteonectina/genética , Animais , Carcinogênese/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Osteonectina/metabolismo , Prognóstico , Transplante Heterólogo , Carga Tumoral/genética
6.
Eur J Med Genet ; 62(7): 103664, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31067494

RESUMO

Ophthalmo-acromelic syndrome is a rare autosomal recessive disorder characterized by ocular and skeletal abnormalities. Ocular findings present as a wide spectrum, ranging from mild microphthalmia to true anophthalmia. Short 5th finger, synostosis of 4th and 5th metacarpals, and oligodactyly in feet are frequent limb malformations. Homozygous variants in the SMOC1 gene (SPARC-related modular calcium-binding protein 1 gene) were identified as causative for the syndrome. A 9-month-old female patient is presented herein, who was diagnosed with ophthalmo-acromelic syndrome and had a homozygous nonsense mutation (p.Arg75Ter) in SMOC1, along with a review of the literature.


Assuntos
Síndrome de Waardenburg/genética , Códon sem Sentido , Feminino , Homozigoto , Humanos , Lactente , Osteonectina/genética , Síndrome de Waardenburg/patologia
7.
PLoS One ; 14(5): e0216934, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31136607

RESUMO

BACKGROUND: Radium-223 is a targeted alpha-particle therapy that improves survival in men with metastatic castration resistant prostate cancer (mCRPC), particularly in men with elevated serum levels of bone alkaline phosphatase (B-ALP). We hypothesized that osteomimicry, a form of epithelial plasticity leading to an osteoblastic phenotype, may contribute to intralesional deposition of radium-223 and subsequent irradiation of the tumor microenvironment. METHODS: We conducted a pharmacodynamic study (NCT02204943) of radium-223 in men with bone mCRPC. Prior to and three and six months after radium-223 treatment initiation, we collected CTCs and metastatic biopsies for phenotypic characterization and CTC genomic analysis. The primary objective was to describe the impact of radium-223 on the prevalence of CTC B-ALP over time. We measured radium-223 decay products in tumor and surrounding normal bone during treatment. We validated genomic findings in a separate independent study of men with bone metastatic mCRPC (n = 45) and publicly accessible data of metastatic CRPC tissues. RESULTS: We enrolled 20 men with symptomatic bone predominant mCRPC and treated with radium-223. We observed greater radium-223 radioactivity levels in metastatic bone tumor containing biopsies compared with adjacent normal bone. We found evidence of persistent Cellsearch CTCs and B-ALP (+) CTCs in the majority of men over time during radium-223 therapy despite serum B-ALP normalization. We identified genomic gains in osteoblast mimicry genes including gains of ALPL, osteopontin, SPARC, OB-cadherin and loss of RUNX2, and validated genomic alterations or increased expression at the DNA and RNA level in an independent cohort of 45 men with bone-metastatic CRPC and in 150 metastatic biopsies from men with mCRPC. CONCLUSIONS: Osteomimicry may contribute in part to the uptake of radium-223 within bone metastases and may thereby enhance the therapeutic benefit of this bone targeting radiotherapy.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Ósseas/radioterapia , Terapia de Alvo Molecular/métodos , Neoplasias de Próstata Resistentes à Castração/radioterapia , Rádio (Elemento)/uso terapêutico , Idoso , Fosfatase Alcalina/sangue , Fosfatase Alcalina/genética , Biomarcadores Tumorais/metabolismo , Biópsia , Neoplasias Ósseas/genética , Neoplasias Ósseas/mortalidade , Neoplasias Ósseas/secundário , Osso e Ossos/metabolismo , Osso e Ossos/patologia , Osso e Ossos/efeitos da radiação , Caderinas/genética , Caderinas/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Progressão da Doença , Expressão Gênica , Humanos , Masculino , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patologia , Células Neoplásicas Circulantes/efeitos da radiação , Osteonectina/genética , Osteonectina/metabolismo , Osteopontina/genética , Osteopontina/metabolismo , Próstata/metabolismo , Próstata/patologia , Próstata/efeitos da radiação , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/mortalidade , Neoplasias de Próstata Resistentes à Castração/patologia , Rádio (Elemento)/sangue , Rádio (Elemento)/farmacocinética , Análise de Sobrevida , Microambiente Tumoral/genética , Microambiente Tumoral/efeitos da radiação
8.
PLoS One ; 14(3): e0214301, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30925184

RESUMO

MicroRNAs (miRNAs) are small noncoding RNAs that critically regulate gene expression. Their abundance and function have been linked to a range of physiologic and pathologic processes. In aged monkey muscle, miR-451a and miR-144-3p were far more abundant than in young monkey muscle. This observation led us to hypothesize that miR-451a and miR-144-3p may influence muscle homeostasis. To test if these conserved microRNAs were implicated in myogenesis, we investigated their function in the mouse myoblast line C2C12. The levels of both microRNAs declined with myogenesis; however, only overexpression of miR-451a, but not miR-144-3p, robustly impeded C2C12 differentiation, suggesting an inhibitory role for miR-451a in myogenesis. Further investigation of the regulatory influence of miR-451a identified as one of the major targets Sparc mRNA, which encodes a secreted protein acidic and rich in cysteine (SPARC) that functions in wound healing and cellular differentiation. In mouse myoblasts, miR-451a suppressed Sparc mRNA translation. Together, our findings indicate that miR-451a is downregulated in differentiated myoblasts and suggest that it decreases C2C12 differentiation at least in part by suppressing SPARC biosynthesis.


Assuntos
MicroRNAs/genética , Desenvolvimento Muscular , Osteonectina/genética , Osteonectina/metabolismo , Regiões 3' não Traduzidas , Animais , Linhagem Celular , Proliferação de Células , Camundongos , Mioblastos/citologia , Mioblastos/metabolismo , Polirribossomos/genética , Polirribossomos/metabolismo , Biossíntese de Proteínas
9.
Clin Transl Oncol ; 21(10): 1327-1335, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30762207

RESUMO

PURPOSE: Glioma is a common malignant tumor of the central nervous system, which is characterized by a low cure rate, high morbidity, and high recurrence rate. Consequently, it is imperative to explore some indicators for prognostic prediction in glioma. METHODS: We obtained glioma data from The Cancer Genome Atlas (TCGA). Differentially expressed genes (DEGs) were obtained by R software from TCGA data sets. Through Cox regression analysis, risk scores were obtained to assess the weighted gene-expression levels, which could predict the prognosis of patients with glioma. The validity and the prognostic value of this model in glioma were confirmed by the manifestation of receiver-operating characteristic (ROC) curves, area under the curve (AUC), and 5-year overall survival (OS). RESULTS: In total, 920 DEGs of transcriptome genes in glioma were extracted from the TCGA database. We identified a novel seven-gene signature associated with glioma. Among them, AL118505.1 and SMOC1 were positively related to the 5-year OS of patients with glioma, showing a better prognosis for glioma; however, RAB42, SHOX2, IGFBP2, HIST1H3G, and IGF2BP3 were negatively related to 5-year OS, displaying a worse prognosis. In addition, according to risk scores, AL118505.1 was also a protective factor, while others were risk factors. Furthermore, the expression levels of SHOX2, IGFBP2, and IGF2BP3 were significantly positively correlated with glioma grades. Receiver-operating characteristic (ROC) curve assessed the accuracy and sensitivity of the gene signature. Each of the seven genes for patients with the distribution of the risk score was presented in the heat map. CONCLUSION: We identified a novel seven-gene signature in patients with glioma, which could be used as a predictor for the prognosis of patients with glioma in the future.


Assuntos
Neoplasias Encefálicas/genética , Expressão Gênica , Genes Neoplásicos , Glioma/genética , Área Sob a Curva , Neoplasias Encefálicas/mortalidade , Glioma/mortalidade , Proteínas de Homeodomínio/genética , Humanos , Proteína 2 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteínas de Neoplasias/genética , Osteonectina/genética , Prognóstico , Proteínas de Ligação a RNA/genética , Curva ROC , Análise de Regressão , Risco , Transcriptoma
10.
Clin Sci (Lond) ; 133(2): 351-365, 2019 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-30626728

RESUMO

Obesity, metabolic syndrome, and type 2 diabetes, three strongly interrelated diseases, are associated to increased morbidity and mortality worldwide. The pathogenesis of obesity-associated disorders is still under study. Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein expressed in many cell types including adipocytes, parenchymal, and non-parenchymal hepatic cells and pancreatic cells. Studies have demonstrated that SPARC inhibits adipogenesis and promotes insulin resistance; in addition, circulating SPARC levels were positively correlated with body mass index in obese individuals. Therefore, SPARC is being proposed as a key factor in the pathogenesis of obesity-associated disorders. The aim of this study is to elucidate the role of SPARC in glucose homeostasis. We show here that SPARC null (SPARC-/-) mice displayed an abnormal insulin-regulated glucose metabolism. SPARC-/- mice presented an increased adipose tissue deposition and an impaired glucose homeostasis as animals aged. In addition, the absence of SPARC worsens high-fat diet-induced diabetes in mice. Interestingly, although SPARC-/- mice on high-fat diet were sensitive to insulin they showed an impaired insulin secretion capacity. Of note, the expression of glucose transporter 2 in islets of SPARC-/- mice was dramatically reduced. The present study provides the first evidence that deleted SPARC expression causes diabetes in mice. Thus, SPARC deficient mice constitute a valuable model for studies concerning obesity and its related metabolic complications, including diabetes.


Assuntos
Glicemia/metabolismo , Diabetes Mellitus Experimental/sangue , Insulina/sangue , Ilhotas Pancreáticas/metabolismo , Osteonectina/metabolismo , Envelhecimento/sangue , Animais , Biomarcadores/sangue , Diabetes Mellitus Experimental/genética , Dieta Hiperlipídica , Sacarose na Dieta , Transportador de Glucose Tipo 2/metabolismo , Homeostase , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteonectina/deficiência , Osteonectina/genética , Via Secretória
11.
Cancer Chemother Pharmacol ; 83(4): 639-647, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30643929

RESUMO

PURPOSE: The purpose of the study is to evaluate the predictive efficacy of secreted protein, acidic and rich in cysteine (SPARC) and the class III ß-tubulin (ß-tubulin III, TUBB3) in predicting therapeutic effect in patients with locally advanced esophageal squamous cell carcinoma(ESCC) who received nab-paclitaxel plus cisplatin neoadjuvant chemotherapy(CT) followed by surgery. METHODS: Patients with stage II to III esophageal squamous cell carcinoma of different stages are recruited. The tumor biopsy tissues prior treatment from enrolled patients were examined by SPARC and TUBB3 immunohistochemistry (IHC). Correlations between SPARC/TUBB3 expression and response to chemotherapy and long-term survival in patients received surgical resection was analyzed. RESULTS: A total of 35 patients with stage II to III esophageal squamous cell carcinoma were enrolled. Of the 35 enrolled patients, 30 successfully completed neoadjuvant chemotherapy and underwent R0 resection, 3 refused surgery after chemotherapy, and 2 failed to undergo radical surgery after chemotherapy. Out of patients undergoing surgery, pathological complete response (pCR) was achieved in 6 patients (6/30, 20%). The 1, 2 and 5-year disease free survival (DFS) rates were 70.0%, 36.6% and 33.3%, respectively. The 1, 2 and 5-year overall survival (OS) rates were 83.3%, 63.3% and 36.6%, respectively. SPARC and TUBB3 IHC was performed on the tumor biopsy tissues which were obtained from patients before treatment. Correlation between SPARC/TUBB3 expression and long-term survival in patients was studied. Both the median DFS and OS between SPARC negative samples and SPARC positive staining samples have no statistical difference. However, the median DFS and OS in TUBB3 negative patients was better than those in TUBB3 positive patients (p = 0.002 for DFS, p = 0.001 for OS). In addition, patients with pCR had longer OS and DFS time than those without pCR.COX regression analysis showed that TUBB3 prior treatment and pCR were independent prognostic factors in ESCC patients undergoing sequential surgery after preoperative chemotherapy. CONCLUSIONS: TUBB3 negative expression prior treatment and pCR may indicate a better prognosis for stage II and III ESCC patients after nab-paclitaxel plus cisplatin neoadjuvant chemotherapy following radical esophagectomy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias Esofágicas/terapia , Carcinoma de Células Escamosas do Esôfago/terapia , Osteonectina/genética , Tubulina (Proteína)/genética , Idoso , Albuminas/administração & dosagem , Cisplatino/administração & dosagem , Intervalo Livre de Doença , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/patologia , Esofagectomia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Terapia Neoadjuvante , Estadiamento de Neoplasias , Paclitaxel/administração & dosagem , Prognóstico , Estudos Prospectivos , Taxa de Sobrevida , Resultado do Tratamento
12.
Eur Surg Res ; 60(1-2): 1-12, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30650425

RESUMO

BACKGROUND: Biliary tract cancers (BTCs) have a poor prognosis. BTCs are characterized by a prominent desmoplastic reaction which possibly contributes to the aggressive phenotype of this tumor. The desmoplastic reaction includes excessive production and deposition of extracellular matrix proteins such as periostin, secreted protein acidic and rich in cysteine (SPARC), thrombospondin-1, as well as accumulation of α-smooth muscle actin-positive cancer-associated fibroblasts and immune cells, secreting growth factors and cytokines including transforming growth factor (TGF)-ß. In the present study, we investigated the expression of SPARC in BTC as well as its possible regulation by TGF-ß. METHODS: Expression levels of Sparc, TGF-ß1 and its receptor ALK5 were evaluated by quantitative real-time PCR in 6 biliary tract cell lines as well as 1 immortalized cholangiocyte cell line (MMNK-1). RNAs from tumor samples of 7 biliary tract cancer patients were analyzed for expression of Sparc, TGF-ß type II receptor (TbRII) as well as Twist and ZO-1. MMNK-1 cells were stimulated with TGF-ß for 24 h, and Sparc, ZO-1 and E-Cadherin expressions were determined. The presence of SPARC protein was analyzed by immunohistochemistry in tumor specimens from 10 patients. RESULTS: When comparing basal Sparc transcript levels in diverse BTC cell lines to MMNK-1 cells, we found that it was strongly downregulated in all cancer cell lines. The remaining expression levels were higher in highly differentiated cell lines (CCSW1, MZChA1, MZChA2 and TFK-1) than in less differentiated and undifferentiated ones (BDC, SKChA1). Expression of Sparc in BTC patient samples showed a significant positive correlation with expression of the epithelial marker ZO-1. In contrast, the mesenchymal marker Twist and the TbRII showed a trend of negative correlation with expression of Sparc in these samples. TGF-ß exposure significantly downregulated Sparc expression in MMNK-1 cholangiocytes in vitro in parallel to downregulation of epithelial markers (E-Cadherin and ZO-1). Finally, SPARC immunostaining was performed in 10 patient samples, and the correlation between absence of SPARC and survival times was analyzed. CONCLUSIONS: These data imply that a decrease in SPARC expression is correlated with dedifferentiation of BTC cells resulting in enhanced EMT being possibly mediated by TGF-ß. Thereby SPARC levels might be a marker for individual prognosis of a patient, and strategies aiming at inhibition of SPARC downregulation might have potential for new future therapies.


Assuntos
Neoplasias do Sistema Biliar/patologia , Transição Epitelial-Mesenquimal , Osteonectina/fisiologia , Diferenciação Celular , Linhagem Celular Tumoral , Regulação para Baixo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Osteonectina/análise , Osteonectina/genética , RNA Mensageiro/análise , Fator de Crescimento Transformador beta/farmacologia , Proteína da Zônula de Oclusão-1/análise
13.
Genetics ; 211(2): 683-702, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30518528

RESUMO

Bone morphogenetic protein (BMP) signaling regulates many different developmental and homeostatic processes in metazoans. The BMP pathway is conserved in Caenorhabditis elegans, and is known to regulate body size and mesoderm development. We have identified the C. elegans smoc-1 (Secreted MOdular Calcium-binding protein-1) gene as a new player in the BMP pathway. smoc-1(0) mutants have a small body size, while overexpression of smoc-1 leads to a long body size and increased expression of the RAD-SMAD (reporter acting downstream of SMAD) BMP reporter, suggesting that SMOC-1 acts as a positive modulator of BMP signaling. Using double-mutant analysis, we showed that SMOC-1 antagonizes the function of the glypican LON-2 and acts through the BMP ligand DBL-1 to regulate BMP signaling. Moreover, SMOC-1 appears to specifically regulate BMP signaling without significant involvement in a TGFß-like pathway that regulates dauer development. We found that smoc-1 is expressed in multiple tissues, including cells of the pharynx, intestine, and posterior hypodermis, and that the expression of smoc-1 in the intestine is positively regulated by BMP signaling. We further established that SMOC-1 functions cell nonautonomously to regulate body size. Human SMOC1 and SMOC2 can each partially rescue the smoc-1(0) mutant phenotype, suggesting that SMOC-1's function in modulating BMP signaling is evolutionarily conserved. Together, our findings highlight a conserved role of SMOC proteins in modulating BMP signaling in metazoans.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Transdução de Sinais , Animais , Tamanho Corporal , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Proteínas de Ligação ao Cálcio/genética , Glipicanas/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Neuropeptídeos/metabolismo , Osteonectina/genética , Osteonectina/metabolismo , Faringe/metabolismo , Fator de Crescimento Transformador beta/metabolismo
14.
Eur J Med Genet ; 62(11): 103578, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30445150

RESUMO

Microphthalmia with limb anomalies (MLA, OMIM, 206920) is a rare autosomal-recessive disease caused by biallelic pathogenic variants in the SMOC1 gene. It is characterized by ocular disorders (microphtalmia or anophtalmia) and limb anomalies (oligodactyly, syndactyly, and synostosis of the 4th and 5th metacarpals), variably associated with long bone hypoplasia, horseshoe kidney, venous anomalies, vertebral anomalies, developmental delay, and intellectual disability. Here, we report the case of a woman who interrupted her pregnancy after ultrasound scans revealed a depression of the frontal bone, posterior fossa anomalies, cerebral ventricular enlargement, cleft spine involving the sacral and lower-lumbar vertebrae, and bilateral microphthalmia. Micrognathia, four fingers in both feet and a slight tibial bowing were added to the clinical picture after fetal autopsy. Exome sequencing identified two variants in the SMOC1 gene, each inherited from one of the parents: c.709G>T - p.(Glu237*) on exon 8 and c.1223G>A - p.(Cys408Tyr) on exon 11, both predicted to be pathogenic by different bioinformatics software. Brain histopathology showed an abnormal cortical neuronal migration, which could be related to the SMOC1 protein function, given its role in cellular signaling, proliferation and migration. Finally, we summarize phenotypic and genetic data of known MLA cases showing that our case has some unique features (Chiari II malformation; focal neuropathological alterations) that could be part of the variable phenotype of SMOC1-associated diseases.


Assuntos
Micrognatismo/genética , Microftalmia/genética , Neurônios/patologia , Osteonectina/genética , Adulto , Alelos , Encéfalo/diagnóstico por imagem , Encéfalo/fisiopatologia , Movimento Celular/genética , Criança , Consanguinidade , Éxons/genética , Feminino , Feto , Homozigoto , Humanos , Lactente , Deformidades Congênitas dos Membros , Masculino , Micrognatismo/diagnóstico , Micrognatismo/diagnóstico por imagem , Micrognatismo/fisiopatologia , Microftalmia/diagnóstico por imagem , Microftalmia/fisiopatologia , Mutação , Linhagem , Análise de Sequência de DNA
15.
Methods Mol Biol ; 1895: 27-42, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30539527

RESUMO

Suicide gene therapy using the herpes simplex virus thymidine kinase (HSV-tk) gene, combined with the prodrug ganciclovir (GCV) medication, is a promising approach for the treatment of malignant tumors, including prostate cancer. The success of this therapeutic strategy requires tissue- or tumor-specific gene expression and efficient gene delivery. In this chapter, we describe the experimental protocols of key methodologies, including promoter construction, reporter assay, adenoviral vector construction and preparation, HSV-tk enzymatic assay and cytotoxicity assay to evaluate the specificity and efficacy of osteonectin promoter-mediated HSV-tk/GCV suicide gene therapy of prostate cancer.


Assuntos
Ganciclovir/metabolismo , Genes Transgênicos Suicidas , Terapia Genética/métodos , Regiões Promotoras Genéticas , Neoplasias da Próstata/terapia , Timidina Quinase/metabolismo , Adenoviridae/genética , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Ganciclovir/uso terapêutico , Vetores Genéticos , Humanos , Masculino , Osteonectina/genética , Pró-Fármacos/metabolismo , Pró-Fármacos/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Simplexvirus/enzimologia , Proteínas Virais/metabolismo
16.
J Gastroenterol Hepatol ; 34(8): 1424-1431, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30422339

RESUMO

BACKGROUND AND AIM: Secreted protein acidic and rich in cysteine (SPARC) is a glycoprotein involved in extracellular matrix remodeling, which regulates cell growth. It could be involved in hepatic fibrogenesis related to chronic inflammations, hepatocellular carcinoma (HCC) angiogenesis, and tumor progression. We aimed to study the expressions of single nucleotide polymorphisms in the SPARC gene and their impact on susceptibility and survival of HCC patients. METHODS: We conducted a case-control study on 200 HCC patients and 50 matched healthy controls. All patients were subjected to laboratory investigations, ultrasound, and real-time polymerase chain reaction to detect the genetic polymorphisms (rs3210714, rs11950384, and rs7719521) in the SPARC gene in the blood. RESULTS: One hundred sixty (80%) patients were men with a mean age of 43 years. The SPARC gene showed a significant higher prevalence of rs3210714 mutation (i.e. AA or AG) and a significant lower prevalence of rs11950384 mutation (i.e. AA or AC) among HCC patients in comparison with controls (83% vs 22%, P ≤ 0.001) and (65.5 vs 86%, P = 0.005), respectively, while rs7719521 mutation did not reach significance. On univariate and multivariate analyses, elder age and having at least one copy of the mutant rs3210714 were associated with a significantly increased risk of HCC (P < 0.001 for both), whereas the presence of at least one copy of the mutant rs11950384 carried a significantly reduced risk of having HCC (P < 0.01). Overall survival did not differ significantly between any of the SPARC gene mutation groups. CONCLUSIONS: The SPARC gene polymorphisms had a diverse impact on the susceptibility of HCC due to its ability to inhibit or promote tumor progression. SPARC gene polymorphisms were not related to survival of our HCC patients, and probably, this needs further analysis of other SPARC gene nucleotides.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Mutação , Osteonectina/genética , Polimorfismo de Nucleotídeo Único , Adulto , Fatores Etários , Idoso , Biomarcadores Tumorais/sangue , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/diagnóstico por imagem , Carcinoma Hepatocelular/mortalidade , Estudos de Casos e Controles , Feminino , Frequência do Gene , Predisposição Genética para Doença , Humanos , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Osteonectina/sangue , Fenótipo , Prognóstico , Medição de Risco , Fatores de Risco , Fatores de Tempo
17.
Cancer Genet ; 228-229: 12-16, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30553465

RESUMO

Aneurysmal bone cyst (ABC) is a benign but locally aggressive, mostly pediatric neoplasm, with characteristic USP6 gene rearrangement that distinguishes it from a secondary ABC and other primary bone tumors. With the advent of next-generation sequencing (NGS) technology, several hitherto unknown USP6 fusion partners have been identified in ABC. Accordingly, we present a case of an 18-year-old male with a solid sub-periosteal primary ABC in the diaphysis of the left femur. Using an NGS-based assay, we identified SPARC-USP6 fusion, which has not previously been described in ABC. Including our case, the list of currently known USP6 fusion partners in primary ABC include: CDH11, CNBP, COL1A1, CTNNB1, EIF1, FOSL2, OMD, PAFAH1B1, RUNX2, SEC31A, SPARC, STAT3 and THRAP3.


Assuntos
Cistos Ósseos Aneurismáticos/genética , Fusão Gênica , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Osteonectina/genética , Proteínas Proto-Oncogênicas/genética , Ubiquitina Tiolesterase/genética , Adolescente , Biópsia , Cistos Ósseos Aneurismáticos/diagnóstico por imagem , Cistos Ósseos Aneurismáticos/patologia , Humanos , Masculino , Reação em Cadeia da Polimerase Via Transcriptase Reversa
18.
Reprod Biol Endocrinol ; 16(1): 116, 2018 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-30424792

RESUMO

BACKGROUND: Matrix metalloproteinases (Mmps) and their tissue inhibitors (Timps) are widely recognized as crucial factors for extracellular matrix remodeling in the ovary and are involved in follicular growth, ovulation, luteinization, and luteolysis during the estrous cycle. Recently, several genes have been associated to the modulation of Mmps activity, including Basigin (Bsg), which induces the expression of Mmps in rat ovaries; Sparc, a TGF-ß modulator that is related to increased expression of Mmps in cancer; and Reck, which is associated with Mmps inhibition. However, the expression pattern of Mmp modulators in ovary dynamics is still largely uncharacterized. METHODS: To characterize the expression pattern of Mmps network members in ovary dynamics, we analyzed the spatio-temporal expression pattern of Reck and Sparc, as well as of Mmp2, Mmp9 and Mmp14 proteins, by immunohistochemistry (IHC), in pre-pubertal rat ovaries obtained from an artificial cycle induced by eCG/hCG, in the different phases of the hormone-induced estrous cycle. We also determined the gene expression profiles of Mmps (2, 9, 13 14), Timps (1, 2, 3), Sparc, Bsg, and Reck to complement this panel. RESULTS: IHC analysis revealed that Mmp protein expression peaks at the early stages of folliculogenesis and ovulation, decreases during ovulation-luteogenesis transition and luteogenesis, increasing again during corpus luteum maintenance and luteolysis. The protein expression patterns of these metalloproteinases and Sparc were inverse relative to the pattern displayed by Reck. We observed that the gene expression peaks of Mmps inhibitors Reck and Timp2 were closely paraleled by Mmp2 and Mmp9 suppression. The opposite was also true: increased Mmp2 and Mmp9 expression was concomitant to reduced Reck and Timp2 levels. CONCLUSION: Therefore, our results generate a spatio-temporal expression profile panel of Mmps and their regulators, suggesting that Reck and Sparc seem to play a role during ovarian dynamics: Reck as a possible inhibitor and Sparc as an inducer of Mmps.


Assuntos
Perfilação da Expressão Gênica , Osteonectina/genética , Ovário/metabolismo , Proteínas Supressoras de Tumor/genética , Animais , Basigina/genética , Basigina/metabolismo , Proteínas Sanguíneas/genética , Proteínas Sanguíneas/metabolismo , Ciclo Estral/genética , Feminino , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Metaloproteinases da Matriz/genética , Metaloproteinases da Matriz/metabolismo , Osteonectina/metabolismo , Ovulação/genética , Ratos Sprague-Dawley , Maturidade Sexual/genética , Fatores de Tempo , Inibidores Teciduais de Metaloproteinases/genética , Inibidores Teciduais de Metaloproteinases/metabolismo , Proteínas Supressoras de Tumor/metabolismo
19.
EBioMedicine ; 38: 238-247, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30470612

RESUMO

BACKGROUND: The genes of SPARC, CCR2, and SMAD3 are implicated in orchestrating inflammatory response that leads to fibrosis in scleroderma and other fibrotic disorders. The aim of the studies is to evaluate synergistic anti-fibrotic potency of the siRNAs of these genes. METHODS: The efficacy of the siRNA-combination was evaluated in bleomycin-induced mouse fibrosis. The pathological changes of skin and lungs of the mice were assessed by hematoxylin and eosin and Masson's trichrome stains. The expression of inflammation and fibrosis associated genes and proteins in the tissues were assessed by real-time RT-PCR, RNA sequencing, Western blots and ELISA. Non-crosslinked fibrillar collagen was measured by the Sircol colorimetric assay. FINDINGS: The applications of the combined siRNAs in bleomycin-induced mice achieved favorable anti-inflammatory and anti-fibrotic effects. Activation of fibroblasts was suppressed in parallel with inhibition of inflammation evidenced by reduced inflammatory cells and proinflammatory cytokines in the BALF and/or the tissues by the treatment. Aberrant expression of the genes normally expressed in fibroblasts, monocytes/ macrophage, endothelial and epithelial cells were significantly restrained after the treatment. In addition, transcriptome profiles indicated that some bleomycin-induced alterations of multiple biological pathways were recovered to varying degrees by the treatment. INTERPRETATION: The application of the combined siRNAs of SPARC, CCR2, and SMAD3 genes ameliorated inflammation and fibrosis in bleomycin-induced mice. It systemically reinstated multiple biopathways, probably through controlling on different cell types including fibroblasts, monocytes/macrophages, endothelial cells and others. The multi-target-combined therapeutic approach examined herein may represent a novel and effective therapy for fibrosis.


Assuntos
Fibrose/genética , Osteonectina/genética , Receptores CCR2/genética , Proteína Smad3/genética , Animais , Biomarcadores , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Fibrose/patologia , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Predisposição Genética para Doença , Inflamação/genética , Inflamação/imunologia , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Camundongos , Osteonectina/metabolismo , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Interferência de RNA , RNA Interferente Pequeno/genética , Receptores CCR2/metabolismo , Pele/metabolismo , Pele/patologia , Proteína Smad3/metabolismo
20.
Biofactors ; 44(5): 453-464, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30346081

RESUMO

Migration and metastasis of tumor cells greatly contributes to the failure of cancer treatment. Recently, the extracellular protein secreted protein acidic and rich in cysteine (SPARC) has been reported closely related to tumorigenesis. Some articles have suggested that SPARC promoted metastasis in several highly metastatic tumors. However, there are also some studies shown that SPARC acted as an antitumor factor. SPARC-induced epithelial-to-mesenchymal transition (EMT) in melanoma cells and promoted EMT in hepatocellular carcinoma. Therefore, the role of SPARC in tumorigenesis and its relationship with EMT is still unclear. In this study, we investigated the expression change of SPARC in A549 and H1299 lung cancer cells undergoing EMT process. Our study indicated that SPARC was upregulated in A549 and H1299 cells EMT process. We further investigated the function of SPARC on proliferation, migration, and EMT process of A549 and H1299 cells. Overexpression of SPARC promoted the migration and EMT of A549 and H1299 cells. Knockdown SPARC inhibited the EMT of A549 cells. Overexpression of SPARC induced the increased expression of p-Akt and P-ERK. Furthermore, exogenous SPARC peptide promoted transforming growth factor (TGF)-ß1-induced EMT of A549 and H1299 cells. SPARC knockdown partially eliminated TGF-ß1 function in inducing EMT of A549 cells. SPARC follistatin-like functional domain reduced the expression of E-cadherin, but had no effect on the expression of p-Akt and p-ERK. In conclusion, we elucidated that SPARC contributes to tumorigenesis by promoting migration and EMT of A549 and H1299 lung cancer cells. These results will provide some new suggestion for lung cancer treatment. © 2018 BioFactors, 44(5):453-464, 2018.


Assuntos
Carcinogênese/genética , Neoplasias Pulmonares/genética , Osteonectina/genética , Fator de Crescimento Transformador beta1/genética , Células A549 , Caderinas/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/genética , Proteína Oncogênica v-akt/genética
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